Electrophoresis Method

ABSTRACT

Provided is an electrophoresis device that, by electrophoresis, feeds a sample into capillaries and optically detects the sample, the electrophoresis device being provided with capillaries, a capillary head provided at the distal end of the capillaries, a phoretic medium-filled container used for electrophoresis and filled with a phoretic medium, a guide member that covers the side surface of the phoretic medium-filled container, a seal member that seals from below the phoretic medium filled in the phoretic medium-filled container, and a plunger that presses the seal member.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. patent application No.16/885,828, filed May 28, 2020, which is a divisional of U.S. PatentApplication No. 15/560,395, filed Sep. 21, 2017, which is a U.S.National Stage of Application No. PCT/JP2015/059532, filed Mar. 27,2015, the disclosures of all of which are expressly incorporated byreference herein.

TECHNICAL FIELD

The present invention relates to an electrophoresis device whichperforms separation analysis of nucleic acid, protein, and the like, andan electrophoresis method thereof.

BACKGROUND ART

Recently, as an electrophoresis device, a capillary electrophoresisdevice in which capillaries are filled with a phoretic medium such aspolymer gel or polymer solution has been widely used.

For example, a capillary electrophoresis device as disclosed in PTL 1has been used from the related art. Since the capillary electrophoresisdevice has higher heat-radiation properties than that of a flat platetype electrophoresis device, and a higher voltage can be applied to asample, there is an advantage that electrophoresis is able to beperformed at high speed. In addition, there are lots of advantages that,for example, a small amount of the sample is enough to complete ananalysis, or automatic filling of the phoretic medium or sampleautomatic injection can be performed, and the capillary electrophoresisdevice is used for various separation analysis measurements as well asanalysis of nucleic acid or protein.

In PTL 1, filling of the phoretic medium is performed using a syringepump. There is a relay flow passage block having a function of thesyringe pump, and filling is performed by connecting capillaries,sucking the phoretic medium using the syringe pump, and discharging thephoretic medium to the capillaries. In addition to the relay flowpassage block, a buffer solution for performing electrophoresis is alsoconnected, and the flow passage is switched by opening and closing avalve inside the relay flow passage block.

In the capillary electrophoresis device, the phoretic medium containeror the capillaries are required to be replaced. However, at the time ofreplacing this component, since a part of the relay flow passage blockis exposed to air, there is a possibility that air is mixed into theflow passage. At the time of performing the electrophoresis, a highvoltage of several to several tens of kV is applied between both ends ofthe flow passage. Therefore, in a case in which bubbles are presentinside the flow passage, there is a possibility that the flow passage iselectrically blocked due to the bubbles. In a case in which the flowpassage is electrically blocked, a high voltage difference occurs at theblocked part so as to cause electric discharge. According to magnitudeof the electric discharge, there is a possibility that the capillaryelectrophoresis device is destroyed. Accordingly, before starting theelectrophoresis, the bubbles need to be removed from the inside of theflow passage.

For example, in a case in which bubbles are present inside the flowpassage of the relay flow passage block, a valve of the flow passageconnected with the buffer solution is opened, and the phoretic medium isallowed to flow to the buffer solution side. Accordingly, the bubblesare removed from a section of flow passage inside the relay flow passageblock. Meanwhile, in a case in which the bubbles are present in the flowpassage of the capillaries, the inside of the capillaries are filledwith the phoretic medium in an amount approximately twice with respectto a volume inside the capillaries. At this time, an inner diameter ofeach of the capillaries is as thin as approximately 50 μm. Therefore,the bubbles flow inside the capillaries with the phoretic medium, andare discharged from the other end of the capillaries. That is, thebubbles can be removed from the insides of the capillaries.

Filling of the phoretic medium to the capillaries can be performed bythe same method as that of the related art. However, in a filling methodof the phoretic medium, the extra phoretic medium is required to removebubbles, such that the extra thereof becomes wasted. In a case in whichanalysis is performed many times at once, since the bubbles are removedonly once, an amount of the wasted phoretic medium is small, but in acase in which the analysis is performed for less times at once, thebubbles are required to be removed whenever the capillaries or thephoretic medium are connected, and the amount of the wasted phoreticmedium per the number of analyses increases. Since the phoretic mediumis expensive, the amount of the wasted phoretic medium increases, andthus running costs thereof increase. In addition, if the electrophoresisis performed in a state in which the bubbles are remained, there is apossibility that the relay flow passage block is damaged. In this case,a damaged part is required to be repaired, and thus it takes a lot oftime until restarting the inspection. Therefore, a user needs to checkthe presence and absence of the bubbles.

PTL 2 illustrates a structure in which bubbles in a relay flow passageblock are not required to be visually checked and a difficulty ofoperating an electrophoresis device is reduced. Specifically, a phoreticmedium container is set to a disposable phoretic medium containerprovided with a liquid feeding mechanism, and capillaries and flowpassages of a phoretic medium and a buffer solution are switched in adetachable manner. Only in a case in which the phoretic medium isfilled, the phoretic medium container and the capillaries are connected,at the time of performing the electrophoresis, the capillaries aredetached from the phoretic medium container, and both ends of each ofthe capillaries are directly soaked into the buffer solution. That is,the relay flow passage block itself is not needed. Accordingly, theamount of the wasted phoretic medium for removing the bubbles isreduced, a flow passage having a risk of mixing of bubbles at the timeof performing the electrophoresis can be removed, and thus the previousvisual checking of bubbles by the user before performing theelectrophoresis can be omitted. Even if the electrophoresis is performedin a state in which the bubbles are remained inside the flow passage,the damaged part can be limited to the capillaries. Also, since thecapillaries are articles of consumption, it does not need to be repairedlike the relay flow passage block. That is, the inspection can berestarted only by replacing the capillaries. Therefore, the timerequired until restarting the inspection can be significantly shortened.

CITATION LIST Patent Literature

PTL 1: JP-A-2008-8621

PTL 2: Japanese Patent No. 5391350

SUMMARY OF INVENTION Technical Problem

As a method in which a liquid feeding mechanism is included in aphoretic medium container as disclosed in PTL 2, for example, there is amethod in which a syringe structure such as a syringe is set to thephoretic medium container and a capillaries are directly connected tothe phoretic medium container. In order to allow the method to bepractically used, the phoretic medium container is required to be madeof an inexpensive resin molded article. Even when the amount of thewasted phoretic medium is reduced, the phoretic medium container becomesexpensive, and thus the running costs thereof cannot be reduced.However, in the method of PTL 2, due to an influence of a liquid feedingpressure of the phoretic medium to the capillaries, stiffness of thephoretic medium container has to increase. If the stiffness of thephoretic medium container increases, the container is difficult to bemade of the resin molded article, and costs of the phoretic mediumcontainer increase.

In addition, in order for the liquid feeding mechanism to be included inthe phoretic medium container, and to detachably connect with thecapillaries, apart connected with the capillaries and a part of theliquid feeding mechanism are necessary to have high pressure resistance.Since the inner diameter of the capillaries are approximately ϕ 50 μmand the phoretic medium having several hundred times higher viscositythan water is injected, it is necessary to apply a pressure of severalMPa thereto.

Further, the remaining amount inside the phoretic medium container ischecked, and the amount of liquid feeding is needed to be managed inmore detail same as that of the related art with resolution performance.If the resolution performance for managing the amount of liquid feedingis crude, the amount of liquid feeding for determining that the insideof the capillaries can be filled with the phoretic medium increases. Inthis way, the phoretic medium for removing the bubbles is not needed,but the amount of the phoretic medium for filling the capillariesincreases, and as a result, the required amount of the phoretic mediumincreases. Accordingly, the running costs cannot be reduced.

In order to solve the above-described problems, it is necessary tosuppress the expansion of the phoretic medium container due to theliquid feeding pressure, with the inexpensive phoretic medium containeras it is. An object of the invention is to provide a capillaryelectrophoresis device in which the problems described above are solved.

Solution to Problem

According to the invention, there is provided an electrophoresis devicewhich feeds a sample into capillaries by electrophoresis and opticallydetects the sample, the device including capillaries, a capillary headthat is provided at a distal end of the capillaries, a phoreticmedium-filled container that is used for the electrophoresis and filledwith a phoretic medium, a guide member that covers a side surface of thephoretic medium-filled container, a seal member that seals the phoreticmedium filled in the phoretic medium-filled container from below, and aplunger that presses the seal member.

ADVANTAGEOUS EFFECTS OF INVENTION

According to the invention, in order to suppress the expansion of thephoretic medium container, a container having high pressure resistancecan be used. In addition, the container can be realized as a phoreticmedium container having a function of feeding of inexpensive liquid.Accordingly, both of reduction in running costs and improvement inworkability of the user can be realized.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 is a schematic view of a configuration of a device of theinvention.

FIG. 2 is a top view of the device of the invention.

FIG. 3 is a sectional view of a device taken along A-A line.

FIG. 4 is a detailed view of a liquid feeding mechanism.

FIG. 5 is a detailed view of a capillary array.

FIG. 6 is a detailed view of an anode side buffer solution container.

FIG. 7 is a detailed view of a cathode side buffer solution.

FIG. 8 is a detailed view of a sample container.

FIG. 9 is a detailed view of a phoretic medium container.

FIG. 10 is a detailed view of installation of the phoretic mediumcontainer.

FIG. 11 is a view of a connection state of the capillary array and acathode side buffer solution container.

FIG. 12 is a view of a connection state of the capillary array and theanode side buffer solution container.

FIG. 13 is a view of a connection state of the capillary array and thephoretic medium container.

FIG. 14 is a workflow of analysis according to this example.

FIG. 15 is a detailed view of a cleaning operation of a capillary head(initial cleaning).

FIG. 16 is a detailed view of a cleaning operation of the capillary head(buffer solution contacting liquid).

FIG. 17 is a detailed view of a cleaning operation of the capillary head(buffer solution cleaning).

FIG. 18 is a detailed view of a liquid feeding operation of a phoreticmedium (initial state).

FIG. 19 is a detailed view of a liquid feeding operation of the phoreticmedium (plunger contact detection).

FIG. 20 is a detailed view of a liquid feeding operation of the phoreticmedium (capillary connection).

FIG. 21 is a detailed view of a liquid feeding operation of the phoreticmedium (phoretic medium injection).

FIG. 22 is a detailed view of a liquid feeding operation of the phoreticmedium (plunger contact release).

FIG. 23 is a detailed view of a liquid feeding operation of the phoreticmedium (residual pressure removing).

FIG. 24 is a detailed view of a liquid feeding operation of the phoreticmedium (capillary connection release).

FIG. 25 is a detailed view of a phoretic medium container according tothe example.

FIG. 26 is a sectional view taken along A-A line according to theexample.

DESCRIPTION OF EMBODIMENTS

In order to suppress the expansion of a phoretic medium container, astructure is produced in which the expansion on a device side where acontainer is set is suppressed without increasing stiffness of thephoretic medium container itself. In addition, a fluid sealed partinside the phoretic medium container has an inner pressure sealingstructure which becomes more sealed when the inner pressure increases.Accordingly, even when an inexpensive phoretic medium container is usedas it is, the expansion thereof is suppressed, and it is possible tomaintain a high pressure resistance. Further, a function of detectingthe remaining amount in the device side of the phoretic mediumcontainer, and a function capable of removing an inner pressure insidethe phoretic medium container after feeding liquid are provided.Accordingly, the remaining amount inside the phoretic medium containerand an amount of feeding liquid can be managed.

Hereinafter, a specific structure will be described.

The phoretic medium container has a syringe structure like a syringe,and a guide component for suppressing expansion of the container itselfis set therein. The guide component has high stiffness, and when thephoretic medium container expands, the container expands until cominginto contact with the guide component, but further expansion thereof issuppressed.

The phoretic medium container and capillaries are in contact with eachother by tying a plurality of the capillaries together, providing acapillary head in which a distal end is sharpened like a needle shape,providing a rubber stopper in the phoretic medium container, and makingthe rubber stopper penetrate through the capillary head. At this time,the rubber stopper presses the capillary head, such that expansion ofthe rubber stopper due to a liquid feeding pressure in the capillaryhead is suppressed.

In the phoretic medium container of a syringe structure, a sealcomponent which can be operated to feed liquid is embedded. A sealsurface of the seal component has a shape and a thickness so as to beeasily deformed due to an inner presser further than a containersyringe. In addition, the seal component is a recessed shape inward thecontainer, a distal end in a recessed shape is set to a seal surface,and thus an inner pressure seal structure which is more sealed isprovided when the inner pressure increases.

The phoretic medium is fed by pressing the seal component of thephoretic medium container from the outside. A liquid feeding mechanism,which is provided with a plunger pressing the seal component, isprovided with an encoder so as to detect a speed change in a case inwhich the plunger is in contact with the seal component. In addition,after the liquid feeding, since a pressure of an inner portion of thephoretic medium container increases, a force of returning the sealcomponent to its original position is activated. The plunger which is incontact with the seal component in this state is separated once.Accordingly, the seal component is moved in a direction of the originalposition, and the inner pressure of the phoretic medium container isremoved.

According to the invention, in order to suppress the expansion of thephoretic medium container, a high pressure resistance container can beused. Further, when a position of the seal component inside the phoreticmedium container is detected, and the remaining voltage inside thephoretic medium container is removed, the remaining amount and theamount of liquid being fed inside the phoretic medium container can bemanaged. In addition, the above description can be realized as aninexpensive phoretic medium container having a liquid feeding function.Accordingly, both reduction of a running cost and improvement ofworkability of a user can be realized.

EXAMPLE 1

Hereinafter, a configuration and arrangement of a device, a maincomponent structure, and an installation method of the invention will bedescribed with reference to FIG. 1 to FIG. 10.

FIG. 1 is a view of a configuration of a capillary electrophoresisdevice to which the invention is applied. The device can be mainlydivided into two units of an auto-sampler unit 150 on a lower portion ofthe device and an irradiation detecting and constant-temperature bath160 on an upper portion of the device.

In the auto-sampler unit 150, a Y axis driving member 85 is mounted on asampler base 80, and driving in a Y axis can be performed. In the Y axisdriving member 85, a Z axis driving member 90 is mounted, and driving ina Z axis can be performed. A sample tray 100 is mounted on the Z axisdriving member 90, and a user sets a phoretic medium container 20, ananode side buffer solution container 30, a cathode side buffer solutioncontainer 40, and a sample container 50 on the sample tray 100. Thesample container 50 is set on an X axis driving member 95 which ismounted on the sample tray 100, and only the sample container 50 on thesample tray 100 can be driven in an X axis. The liquid feeding mechanism60 is also provided in the Z axis driving member 90. The liquid feedingmechanism 60 is disposed under the phoretic medium container 20.

The constant-temperature bath 110 and a constant-temperature bath door120 are provided in the irradiation detecting and constant-temperaturebath 160, and a temperature inside the bath can be constantlymaintained. An irradiation detecting unit 130 is mounted on a rear sideof the constant-temperature bath 110, such that detection at the time ofelectrophoresis can be performed. The user sets the capillary array 10in the constant-temperature bath 110, the electrophoresis is performedwhile the capillary array 10 is maintained at a constant temperature inthe constant-temperature bath 110, and the irradiation detecting unit130 performs detection. In addition, an electrode 115, which is fordropping a voltage to GND at the time of applying a high voltage forperforming the electrophoresis, is also mounted in theconstant-temperature bath 110.

As described above, the capillary array 10 is fixed to theconstant-temperature bath 110. The phoretic medium container 20, theanode side buffer solution container 30, the cathode side buffersolution container 40, and the sample container 50 can be driven in a YZaxis by the auto-sampler unit 150, and only the sample container 50 canbe further driven in the X axis. With the fixed capillary array 10, thephoretic medium container 20, the anode side buffer solution container30, the cathode side buffer solution container 40, and the samplecontainer 50 can be automatically connected to an arbitrary position dueto movement of the auto-sampler unit 150.

FIG. 2 is a view of the capillary electrophoresis device when seen fromthe top. An anode side cleaning layer 31, a buffer solution layer foranode side electrophoresis 32, and a buffer solution layer for sampleintroduction 33 are provided in the anode side buffer solution container30 set on the sample tray 100. In addition, a waste liquid layer 41, acathode side cleaning layer 42, and a buffer solution layer for cathodeside electrophoresis 43 are provided in the cathode side buffer solutioncontainer 40.

The phoretic medium container 20, the anode side buffer solutioncontainer 30, the cathode side buffer solution container 40, and thesample container 50 are disposed with a positional relationship asillustrated in drawings. Accordingly, a positional relationship betweenthe anode side and the cathode side at the time of connecting to thecapillary array 10 becomes a positional relationship of “the phoreticmedium container 20 and the waste liquid layer 41”, “the anode sidecleaning layer 31 and the cathode side cleaning layer 42”, “the buffersolution layer for anode side electrophoresis 32 and the buffer solutionlayer for cathode side electrophoresis 43”, and “the buffer solutionlayer for sample introduction 33 and the sample container 50”.

FIG. 3 illustrates a sectional view taken along A-A line of FIG. 2. Thephoretic medium container 20 is set to be inserted into a guide 101which is embedded in the sample tray 100. In addition, the liquidfeeding mechanism 60 is disposed so that a plunger 61 embedded in theliquid feeding mechanism 60 is positioned under the phoretic mediumcontainer 20.

At the time of the electrophoresis, the right side in FIG. 3 of thecapillary array 10 is set as a cathode side, and the left side thereofis set as an anode side. The auto-sampler unit 150 is moved to aposition relating to the positional relationship of “the buffer solutionlayer for anode side electrophoresis 32 and the buffer solution layerfor cathode side electrophoresis 43”, and a high voltage is applied tothe capillary array 10 of the cathode side and flows to GND to theelectrode 115 through the cathode side buffer solution container 40 andthe anode side buffer solution container 30, such that theelectrophoresis is performed.

FIG. 4 illustrates a detailed view of the liquid feeding mechanism 60. Astepping motor 62 attached to a rotary encoder 63 is mounted on a liquidfeeding mechanism base 70, and a driving pulley 67 is attached to thestepping motor 62. For example, the stepping motor 62 is a two-phasestepping motor, and the rotary encoder 63 is capable of performing 400counts per one rotation. A driving pulley 67 and a driven pulley 68 areconnected by a belt 69, and the driven pulley 68 and a ball screw 65 arefixed. A linear guide 66 is attached to the liquid feeding mechanismbase 70 in parallel to the ball screw 65, and the linear guide 66 andthe ball screw 65 are fixed by the slider 71. A detection plate 72 isattached to the slider 71, and an origin point is detected by shieldingan origin sensor 64 using the detection plate 72. In addition, theplunger 61 is attached to the slider 71 in an axis direction same asthat of a driving axis. Accordingly, the plunger 61 can be driven byrotating the stepping motor 62.

FIG. 5 is a detailed view of the capillary array 10. Capillaries 11which are glass tubes having approximately ϕ 50 μm of an inner diameterare provided in the capillary array 10, and a detecting unit 12 isattached to the capillaries 11. The irradiation detecting unit 130detects the detecting unit 12. A load header 16 and SUS pipes 17 areattached to end portions of the cathode sides of the capillaries 11. Asa material of the load header 16, for example, a PBT resin or the likeis desirable which is a resin having a high insulating property and ahigh comparison tracking index. A component which attains conducting ofall of the SUS pipes 17 is embedded in the load header 16, and a highvoltage is applied to all the SUS pipes 17 by applying the high voltageto the component. Each of the capillaries 11 penetrates and is fixed tothe SUS pipes 17. On the anode side, a plurality of the capillaries 11are tied together by the capillary head 13. The capillary head 13includes a capillary head distal end 15 which is a needle shape at anacute angle, and a capillary head boss 14 which is a part having abigger outer diameter than that of the capillary head distal end 15. Asa material of the capillary head 13, a PEEK resin or the like, which hasstiffness so as to be hardly broken and is highly stable againstchemicals and analysis, is desirable.

It is omitted in the drawings, but the detecting unit 12, the loadheader 16, and the capillary head 13 are respectively fixed, when thecapillary array 10 is fixed to the constant-temperature bath 110. Aposition of the detecting unit 12 is determined with high accuracy so asto be detected by the irradiation detecting unit. The load header 16 isfixed so as to be conducted with a part to which a high voltage isapplied at the time of being fixed. The capillary head 13 is positionedso that the capillary head distal end 15 is directly positioned underthe capillary head, and is strongly fixed so as to withstand a load.Regarding the positional relationship of the cathode side and the anodeside at the time of fixing, the plurality of capillaries 11 are disposedso as not to overlap with each other at the time of being set in adevice.

FIG. 6 is a detailed view of the anode side buffer solution container30. As described above, the anode side cleaning layer 31, the buffersolution layer for anode side electrophoresis 32, and the buffersolution layer for sample introduction 33 are provided in the anode sidebuffer solution container 30. One container is partitioned by apartition 56. As a material of the anode side buffer solution container30, a PC resin or the like which is a transparent resin in order to beable to see the buffer solution therein is desirable. Sections of theanode side cleaning layer 31, the buffer solution layer for anode sideelectrophoresis 32, and the buffer solution layer for sampleintroduction 33 are respectively illustrated as a sectional view of B-B,a sectional view of C-C, and a sectional view of D-D. As illustrated ineach sectional view, an upper surface of the anode side buffer solutioncontainer 30 is sealed with a film 55. As a material of the film 55, amaterial is used which is able to weld with a PC resin and to suppresswater vapor permeation. In addition, a cleaning operation to bedescribed later includes an expanding operation of a hole. When theexpanding operation is considered, since the film 55 is required to bemade with a material which is difficult to stretch, it is desirable thatthe film includes an aluminum layer. Among them, an anode side cleaningliquid 35, a buffer solution for anode side electrophoresis 36, and abuffer solution for sample introduction 37 are respectively enclosed.Respective liquid is enclosed in an amount with which the analysis canbe performed 10 times. Since an upper portion of the partition 56 isalso sealed with the film 55, reagents are not mixed with each other. Inaddition, the container bottom of the anode side cleaning layer 31 withrespect to the buffer solution layer for anode side electrophoresis 32and the buffer solution layer for sample introduction 33 is deep. It isomitted in the drawings, but when the anode side buffer solutioncontainer 30 is set in the device, the anode side buffer solutioncontainer is set in the device as it is without peeling off the film 55,and is locked so as not to be lifted.

FIG. 7 illustrates a detailed view of the cathode side buffer solutioncontainer 40. As described above, the waste liquid layer 41, the cathodeside cleaning layer 42, and the buffer solution layer for cathode sideelectrophoresis 43 are provided in the cathode side buffer solutioncontainer 40. These components become one container, and it ispartitioned by a partition 56. A material of the cathode side buffersolution container 40 is desirably a PC resin or the like, which is atransparent resin in order to be possible to see the buffer solutiontherein, same as the material of the anode side buffer solutioncontainer 30. Sections of the waste liquid layer 41, the cathode sidecleaning layer 42, and the buffer solution layer for cathode sideelectrophoresis 43 are respectively illustrated as a sectional view ofE-E, a sectional view of F-F, and a sectional view of G-G. Asillustrated in each sectional view, an upper surface of the cathode sidebuffer solution container 40 is sealed with the film 55, same as theanode side buffer solution container 30. All layers have the same shape,and a waste liquid received liquid 45, a cathode side cleaning liquid46, and a buffer solution for cathode side electrophoresis 47 areenclosed therein. Respective liquid is enclosed in an amount with whichthe analysis can be performed 10 times. Since the upper portion of thepartition 56 is also sealed with the film 55 same as the anode sidebuffer solution container 30, reagents are not mixed with each other. Itis omitted in the drawings, but when the cathode side buffer solutioncontainer 40 is set in the device, the cathode side buffer solutioncontainer is set in the device as it is without peeling the film 55, andlocked so as not to be lifted.

FIG. 8 illustrates a detailed view of the sample container 50. Asectional view of H-H is set to a sectional view of the sample container50, and a sample 51 is enclosed in the sample container. Since a userprepares the sample 51, the sample container 50 is preferably acontainer which is also easily handled in a pretreatment process or thelike. In this device, the sample container 50 is for example, anEppendorf's eight strips tube. It is omitted in the drawings, but whenthe sample container 50 is set in the device, the sample container isset in the device as it is.

FIG. 9 illustrates a detailed view of the phoretic medium container 20.In the phoretic medium container 20, a seal 22 of the recessed shape isembedded in the syringe 21, and the container is sealed with a cap 24 byputting a rubber stopper 23 from the top. An upper portion of the cap 24is further sealed with the film 55. A material of the syringe 21 isdesirably a PP resin or the like which is a resin capable of beingthinly deformed. A material of the seal 22 is desirably an ultra highmolecular PE resin or the like, which is frequently used for sealingliquid of a sliding portion and have excellent sliding properties. Amaterial of the rubber stopper 23 is desirably a silicon rubber or thelike which is stable with respect to analysis. A material of the cap 24is desirably a PC resin or the like in order to be uniform with the film55 of each container. The phoretic medium 26 is enclosed therein, andair 27 which has entered at the time of enclosing is enclosed so as tobe accumulated at an upper portion. The phoretic medium 26 is enclosedin an amount with which the analysis can be performed 10 times. When theseal 22 is applied with a load from the outside, an inner portion of thesyringe 21 can be operated.

FIG. 10 illustrates a detailed view of installation of the phoreticmedium container 20. When the phoretic medium container 20 is set in thedevice, first, the film 55 attached to the cap 24 is peeled off. Afterthat, the cap is inserted into a guide 101 embedded in the sample tray100, and is fixed from the top so as not to be lifted. At this time, agap between an outer diameter of the syringe 21 and an inner diameter ofthe guide 101 is made as small as possible. The smaller the gap is, thebetter, but the outer diameter of the syringe 21 of a resin moldedarticle and the inner diameter of the guide 101 which is a machinedarticle are set as a gap which is reasonable for processing.Specifically, the gap is approximately 0.1 mm.

Hereinafter, with reference to FIG. 11 to FIG. 13, a connection methodof the capillary array 10 and the phoretic medium container 20, theanode side buffer solution container 30, and the cathode side buffersolution container 40 in the invention will be described.

FIG. 11 illustrates a connection state of the capillary array 10 and thecathode side buffer solution container 40. The cathode side buffersolution container 40 set in the sample tray 100 is connected to thefixed capillary array 10 by the Z axis driving of the auto-sampler unit150. At the time of connecting, the film 55 is connected to anillustrated position by penetrating the SUS pipes 17. This connectionmethod is the same as that of all of the waste liquid layer 41, thecathode side cleaning layer 42, and the buffer solution layer forcathode side electrophoresis 43.

FIG. 12 illustrates a connection state of the capillary array 10 and theanode side buffer solution container 30. The anode side buffer solutioncontainer 30 set in the sample tray 100 is connected to the fixedcapillary array 10 by the Z axis driving of the auto-sampler unit 150.At the time of connecting, the film 55 is connected to an illustratedposition by penetrating the capillary head 13 and the electrode 115.This connection method is the same as that of all of the anode sidecleaning layer 31, the buffer solution layer for anode sideelectrophoresis 32, and the buffer solution layer for sampleintroduction 33, but only a depth of the anode side cleaning layer 31 tobe inserted is changed.

When the anode side buffer solution container 30 and the cathode sidebuffer solution container 40 are connected, these containers may beconnected by peeling off the film 55 without penetrating the film. Inthis manner, load with respect to the SUS pipes 17 or the capillary head13 disappears, but when the cathode side buffer solution container 40 isset in the sample tray 100, the buffer solution or the cleaning liquidis able to be spilled, and the buffer solution or the cleaning liquidevaporates during analysis. Here, the upper portion of the containermaybe covered with not only the film 55 but also a rubber scepter withincision. Accordingly, the buffer solution or the cleaning liquid doesnot spill and is prevented from evaporating, thereby making it possibleto reduce the load on the SUS pipes 17 or the capillary head 13.

FIG. 13 illustrates a connection state of the capillary array 10 and thephoretic medium container 20. The phoretic medium container 20 set inthe sample tray 100 is connected to the fixed capillary array 10 by theZ axis driving of the auto-sampler unit 150. At the time of connecting,the container is connected by making the capillary head 13 penetrate therubber stopper 23. Since the capillary head distal end 15 is a needleshape, the capillary head distal end is also capable of penetrating therubber stopper 23. At this time, the electrode 115 has a positionalrelationship of not being in contact with the phoretic medium container20. The capillary head 13 includes the capillary head boss 14 in whichthe outer diameter is great, and the capillary head is connected to thecapillary head boss 14 by pressing down the upper surface of the rubberstopper 23 from the top. In addition, the air 27 also enters the upperportion inside the phoretic medium container 20, but the capillary headdistal end 15 after being inserted is disposed so as to be positionedunder the air 27.

At this time, the film 55 of the phoretic medium container 20 is peeledoff and set, but the film 55 may penetrate the capillary head 13 bysetting the film 55 without being peeled. Accordingly, the load on thecapillary head 13 increases, but it is also possible to preventforgetting to peel off the film 55, and workability of the user isimproved.

Hereinafter, a workflow of analysis in the example will be describedwith reference to FIG. 14.

In Step 200, the user sets the capillary array 10 in theconstant-temperature bath 110. In addition, the phoretic mediumcontainer 20, the anode side buffer solution container 30, the cathodeside buffer solution container 40, and the sample container 50 are setin the sample tray 100. It is omitted in the drawings, but a barcode isattached to the capillary array 10, the phoretic medium container 20,the anode side buffer solution container 30, the cathode side buffersolution container 40, and the sample container 50, which are articlesof consumption. When each of the articles of consumption is set in thedevice, the user reads information relating to the barcode of each ofthe articles of consumption using a barcode leader mounted in thedevice. Accordingly, a serial number, an expiration date, the number oftimes of use, or the like of each article of consumption can be managed.

In Step 201, the set capillary array 10 is maintained at a constanttemperature by the constant-temperature bath 110.

In Step 202, due to movement of Y axis driving and Z axis driving of theauto-sampler unit 150, the capillary head 13 and the SUS pipes 17 of thecapillary array 10 are respectively inserted into the anode sidecleaning layer 31 and the cathode side cleaning layer 42. Accordingly,the capillary head 13 and the SUS pipes 17 are cleaned. The cleaningoperation of the capillary head 13 side will be described later indetail with reference to FIG. 15 to FIG. 17.

In Step 203, due to movement of Y axis driving and Z axis driving of theauto-sampler unit 150, the capillary head 13 and the SUS pipes 17 of thecapillary array 10 are respectively inserted into the phoretic mediumcontainer 20 and the waste liquid layer 41. In this state, the liquidfeeding mechanism 60 is driven, and the phoretic medium 26 enclosed inthe phoretic medium container 20 is fed to the capillaries 11. Theliquid feeding operation will be described later in detail withreference to FIG. 18 to FIG. 24.

In Step 202, due to movement of Y axis driving and Z axis driving of theauto-sampler unit 150 again, the capillary head 13 and the SUS pipes 17of the capillary array 10 are respectively inserted into the anode sidecleaning layer 31 and the cathode side cleaning layer 42. Accordingly,the capillary head 13 and the SUS pipes 17 are cleaned.

In Step 204, due to movement of Y axis driving and Z axis driving of theauto-sampler unit 150, the capillary head 13 and the SUS pipes 17 of thecapillary array 10 are respectively inserted into the buffer solutionlayer for sample introduction 33 and the sample container 50. At thistime, the electrode 115 is also inserted into the buffer solution layerfor sample introduction 33. Accordingly, both ends of each of thecapillaries 11 are conducted. A high voltage is applied in this state,and the sample 51 is inserted into the capillaries 11.

In Step 202, due to movement of Y axis driving and Z axis driving of theauto-sampler unit 150 again, the capillary head 13 and the SUS pipes 17of the capillary array 10 are respectively inserted into the anode sidecleaning layer 31 and the cathode side cleaning layer 42. Accordingly,the capillary head 13 and the SUS pipes 17 are cleaned.

In Step 205, due to movement of Y axis driving and Z axis driving of theauto-sampler unit 150 again, the capillary head 13 and the SUS pipes 17of the capillary array 10 are respectively inserted into the buffersolution layer for anode side electrophoresis 32 and the buffer solutionlayer for cathode side electrophoresis 43. At this time, the electrode115 is also inserted into the buffer solution layer for sampleintroduction 33. Accordingly, both ends of each of the capillaries 11are conducted. A high voltage is applied in this state, andelectrophoresis is performed. The sample 51 on which the electrophoresisis performed is detected by the irradiation detecting unit 130.

In Step 202, due to movement of Y axis driving and Z axis driving of theauto-sampler unit 150 again, the capillary head 13 and the SUS pipes 17of the capillary array 10 are respectively inserted into the anode sidecleaning layer 31 and the cathode side cleaning layer 42. Accordingly,the capillary head 13 and the SUS pipes 17 are cleaned.

One analysis is finished by analyzing data detected due to this seriesof movements. In a case in which the analysis is continuously performed,the X driving member 95 on the sample tray 100 is driven, and theposition of the sample container 50 is switched, such that theabove-described operation is repeated.

Hereinafter, the cleaning operation of the capillary head 13 will bedescribed in detail with reference to FIG. 15 to FIG. 17. The capillaryhead 13 is associated with contacting liquid with the phoretic medium 26during the analysis. Ina case in which the analysis is continuouslyperformed, the buffer solution enters into the phoretic medium 26 at thetime of contacting liquid. If the phoretic medium 26 in which the buffersolution is mixed is fed to the capillaries 11 as it is, the analysisproperties deteriorate. In addition, the phoretic medium 26 hasproperties of crystallizing when dried. If the phoretic medium remainsas being crystallized, the capillaries 11 are clogged when the liquid isfed, and there is a concern that the phoretic medium 26 cannot be fed.Further, the phoretic medium 26, which is crystallized, is inserted intoa connection portion of the capillary head 13 and the phoretic mediumcontainer 20, and there is a concern that the phoretic medium 26 leaksat the time of feeding the liquid. Therefore, cleaning of the capillaryhead 13 is very important.

FIG. 15 illustrates initial cleaning in an analysis workflow in detail.The anode side cleaning layer 31 of the anode side buffer solutioncontainer 30 is inserted into the capillary head 13 and the electrode115. At this time, as illustrated, the capillary head boss 14 part isinserted into the film 55 until a hole is made. After that, the anodeside cleaning layer 31 is pulled out. The outer diameter of thecapillary head boss 14 is greater than the outer diameter of thecapillary head distal end 15, and an extension hole 150 greater than thecapillary head distal end 15 is made to be empty on the film 55.

FIG. 16 illustrates a state of contacting liquid with the buffersolution in the analysis workflow in detail. The buffer solution layerfor sample introduction 33 or the buffer solution layer for anode sideelectrophoresis 32 of the anode side buffer solution container 30 isinserted into the capillary head 13 and the electrode 115. At this time,as illustrated, insertion depths of these components are more shallowthan a depth of a case of being inserted into an anode side cleaningliquid layer 31. Accordingly, a penetrating hole 151 is opened to thefilm 55, and the buffer solution adhered to the capillary head 13 is ina range of a buffer solution adhering range 155. It is omitted in thedrawings, but the phoretic medium container 20 at the time of beingconnected is connected, so that a range of the phoretic medium 26adhered to the capillary head 13 is the same as the range describedabove.

FIG. 17 illustrates cleaning in the analysis workflow in detail. Afterinitial cleaning is performed on the capillary head 13 and the electrode115 once, the anode side cleaning layer 31 of the anode side buffersolution container 30 is inserted thereto. As illustrated, the extensionhole 150 is empty on the film 55 of the anode side cleaning layer 31.Since the extension hole 150 is greater than the outer diameter of thecapillary head distal end 15, the capillary head distal end 15 does notcome into contact with the film 55. Therefore, a part of the buffersolution adhering range 155 adhered to the capillary head distal end 15does not come into contact with the film 55, and can be cleaned by theanode side cleaning liquid 35. In addition, since the insertion depth isalso deep, the cleaning can be performed up to a cleaning range 156, andthe entire part of the buffer solution adhering range 155 adhered to thecapillary head 13 can be cleaned.

The cleaning is performed by this series of operations, but speed at thetime of pulling out each solution capillary head 13 slows downextremely. Accordingly, an original amount of solution to be adhered tothe capillary head 13 is reduced, and a carrying amount of the solutionis reduced.

In a case in which efficiency of the cleaning is desired to beincreased, two of the anode side cleaning layers 31 may be provided. Inaddition, the upper portion of the container may be covered not with thefilm 55 but with a rubber scepter with incision, and a method of wipingthe solution adhered to the capillary head 13 may be used. If the numberof continuously performing analysis is small, an amount of the buffersolution mixed with the phoretic medium 26 is reduced. In this case, thecleaning operation itself is not required.

Hereinafter, the liquid feeding operation of the phoretic medium 26 willbe described in detail with reference to FIG. 18 to FIG. 24.

FIG. 18 illustrates a view of an initial state, which is a series ofmovements of an injecting operation of the phoretic medium 26. Asdescribed above, the phoretic medium container 20 is set by beinginserted into the guide 101 embedded in the sample tray 100. At thistime, the plunger 61 of the liquid feeding mechanism 60 is disposeddirectly under the phoretic medium container 20, and the seal 22 insidethe phoretic medium container 20 can be operated by moving the plunger61.

FIG. 19 illustrates a view of a contact detection state of the plunger61, which is a series of movements of the injecting operations of thephoretic medium 26. First, as illustrated in FIG. 18, the plunger 61 ofthe liquid feeding mechanism 60 is brought into contact with the seal 22inside the phoretic medium container 20, so that a position thereof isdetected. The stepping motor 62 of the liquid feeding mechanism 60 isdriven with a weak driving current, and the stepping motor 62 is steppedout at the time of coming into contact with the seal 22. In order toreduce the load on the seal 22, the driving current of the steppingmotor 62 is adjusted so that a thrust force of the plunger 61 at thistime is approximately 10 N. The contact of the plunger 61 is detected bydetecting stepping-out of the stepping motor 62 at this time with therotary encoder 63. When the contact position of the plunger 61 isdetected, an amount of the phoretic medium 26 inside the phoretic mediumcontainer 20 is accurately checked so as to be capable of being used formanaging an amount of liquid being fed or detecting liquid leaking.After the contact of the plunger 61 is detected, the plunger 61 isexcited with a larger current than a current at the time of driving, andis held at a position where the seal 22 is in contact. A current valueat the time of exciting is desirably set to a current value which iscapable of maintaining the thrust force equal to a pressure generated atthe time of feeding the phoretic medium 21.

FIG. 20 illustrates a view of a connection state of the capillary head13, which is a series of movements of the injecting operations of thephoretic medium 26. Due to the movement of the Z axis driving member 90of the auto-sampler unit 150, the capillary head 13 and the phoreticmedium container 20 are connected. As described above, the sharpcapillary head distal end 15 penetrates through and is connected to therubber stopper 23 inside the phoretic medium container 20. Since theplunger 61 of the liquid feeding mechanism 60 is mounted on the Zdriving member 90 of the auto-sampler unit 150, the plunger 61 isconnected to the seal 22 in a state of being in contact. In addition, asdescribed above, the plunger 61 is connected to the capillary head boss14 by pressing down the upper surface of the rubber stopper 23 from thetop. At this time, the capillary head 13 is inserted into the phoreticmedium container 20 by the rubber stopper 23 in a state of being sealed.Accordingly, a volume change inside the phoretic medium container 20occurs, and a pressure inside the phoretic medium container 20increases; however, since the plunger 61 presses the seal 22 down, theseal 22 is not operated.

FIG. 21 illustrates a view of an injection state of the phoretic medium26, which is a series of movements of the injecting operations of thephoretic medium 26. After the connection of the capillary head 13, theseal 22 is operated by driving the plunger 61 with the liquid feedingmechanism 60, the volume inside the phoretic medium container 20 ischanged, such that the liquid is fed. At this time, the inside of thephoretic medium container 20 is highly pressurized, and each componentof the phoretic medium container 20 expands. Since the phoretic mediumcontainer 20 at this time has low stiffness, the amount of expansionthereof is great, and the container becomes unstable. Therefore, theexpansion of the phoretic medium container 20 causes great influence onsealing properties of the phoretic medium 26.

Here, the guide 101 suppresses the expansion of the syringe 21. Inaddition, the capillary head 13 suppresses the expansion of the rubberstopper 23. Further, since a shape of the seal 22 is a recessed shape,when the seal 22 expands due to the inner pressure, the inside thereofbecomes more sealed. The seal 22 is formed in a shape or a stiffnesseasy to expand more than the syringe 21, and the influence due to theexpansion of the syringe 21 can be reduced. Specifically, a thickness ofthe syringe 21 is set to 1 mm, and a thickness of the seal 22 is set toapproximately 0.6 mm, such that there are differences in expansionfactors.

Accordingly, influence due to the expansion on the sealing properties isreduced. However, no matter how much the expansion amount reduces, theexpansion amount cannot be removed. The expansion amount is varied so asto affect the management of an amount of liquid feeding.

Here, first, the stepping motor 62 is driven by a driving current whichis a pressure required to feed liquid, and the plunger 61 is driven. Apressure required for feeding liquid at this time is set to 3 MPa, andin order to generate the pressure, the driving current of the steppingmotor 62 is adjusted so that a thrust force of the plunger 61 is set to75 N. Accordingly, the inside of the phoretic medium container 20expands, but the stepping motor 62 performs stepping-out when the innerpressure is increased as much as needed. At this time, since thephoretic medium container 20 becomes expanded, the stepping-out isdetected by the rotary encoder 63. Even after the stepping-out isdetected, the stepping motor 62 performs stepping-out and iscontinuously driven. Since the phoretic medium 26 is gradually fed intothe capillaries 11, the plunger 61 is gradually driven. Also, afterdetecting that the phoretic medium container 20 expanded, an amountbeing driven by the plunger 61 is detected by the rotary encoder 63, andthe amount as much as required for the phoretic medium 26 is fed to thecapillaries 11. With such a liquid feeding method, the amount of liquidfeeding can be managed without receiving an influence due to theexpansion of the phoretic medium container 20.

FIG. 22 and FIG. 23 illustrate a view of an operation of removing aresidual pressure inside the phoretic medium container 20 in detail,which is a series of movements of the injecting operations of thephoretic medium 26. After the liquid feeding is completed, the plunger61 of the liquid feeding mechanism 60 is dropped as illustrated in FIG.21, and the contact with the seal 22 is released. After the liquidfeeding is completed, the pressure inside the phoretic medium container20 remains high. However, according to this operation, as illustrated inFIG. 22, the seal 22 is pressed back due to the pressure inside thephoretic medium container 20, and the residual pressure inside thephoretic medium container 20 is removed.

FIG. 24 illustrates a view of a connection releasing operation of thecapillary head 13 in detail, which is a series of movements of theinjecting operations of the phoretic medium 26. Due to the movement ofthe Z axis driving member 90 of the auto-sampler unit 150, connection ofthe capillary head 13 and the phoretic medium container 20 is released.At this time, since the residual pressure inside the phoretic mediumcontainer 20 is removed by previous operation, there is no need toconsider spattering of the phoretic medium 26 at the time of releasingthe connection of the capillary head 13 and the phoretic mediumcontainer 20. By the operation described above, the phoretic medium 26is fed to the capillaries 11.

When applying this structure and the liquid feeding operation describedabove, the expansion of the phoretic medium container 20 is suppressedand influence due to the expansion is reduced, and thereby making itpossible to manage the amount of liquid feeding. In addition, theabove-described aspect can be realized as an inexpensive phoretic mediumcontainer having a function of liquid feeding. Accordingly, bothreduction of running costs and improvement of workability of the usercan be realized.

Example 2

When contacting of the plunger 61 at the time of the liquid feedingoperation is detected, a separate sensor maybe provided. If the plunger61 is in contact with the seal 22, a spring is bent, and the detectionplate may partition the sensor by the bending. Otherwise, a contact typeswitch may be mounted.

Example 3

When residual pressure is removed at the time of the liquid feedingoperation, as sliding resistance of the seal 22, the residual pressureinside the phoretic medium container 20 is remained. Here, the plunger61 and the seal 22 are fixed, and the residual pressure may be removedby forcedly lowering the seal 22. At this time, a mechanism for fixingthe plunger 61 and the seal 22 is needed, but the residual pressure dueto sliding resistance of the seal 22 can be also removed.

Example 4

FIG. 25 illustrates another aspect of the phoretic medium container 20.In FIG. 25, the aspect thereof is the phoretic medium container 20 inwhich a bottom of the syringe 21 is sealed and the rubber stopper 23having a function same as that of the seal 22 is embedded. Due to onlythe movement of the Z axis driving member 90 of the auto-sampler unit150, the capillary head 13 penetrates the rubber stopper 23 of thephoretic medium container 20, the capillary head 13 is further pushed,and thus the phoretic medium 26 can be also fed to the capillaries 11.At this time, a force for penetrating the rubber stopper 23 to thecapillary head 11 is greater than a sliding resistance at the time offeeding liquid. When this aspect is applied, the liquid feedingmechanism 60 itself can be also eliminated.

Example 5

As illustrated in FIG. 26, the auto-sampler unit 150 may be divided intoan anode side and a cathode side. In addition, the X axis driving member95 for switching the sample is mounted on the sampler base 80 but not onthe sample tray 100, and may be driven in an X axis of each sample tray100. Further, the electrode 115 of the anode side may be integrated withthe capillary head 13. The capillary head 13 is made of a materialcapable of being electrically conducted, and a voltage is dropped to GNDat the time of fixing the capillary head 13, and thus the electrode 115itself is not needed.

Example 6

A plurality of the phoretic medium containers 20 maybe set. When theplurality of phoretic medium containers are set, it is possible toincrease the number of continuously performing RUN. Otherwise, accordingto the number of continuously performing RUN, an amount of the phoreticmedium 26 enclosed in the phoretic medium container 20 may be varied. Inaddition, since the type of the phoretic medium container 20 is set toone, the phoretic medium 26 as 1 RUN is enclosed in the phoretic mediumcontainer 20, and the phoretic medium container may be set as much asthe number of continuously performing RUN.

Example 7

The type of the phoretic medium 26 enclosed in the phoretic mediumcontainer 20 may be pluralized. There are multiple types of the phoreticmediums 26, and the appropriate phoretic medium 26 is differentdepending on contents of the analysis. Here, when the structure of thephoretic medium container 20 is set to be the same, and the types of thephoretic medium 26 enclosed therein are changed, it is possible tocorrespond to various analyses. The management at this time is performedusing barcodes attached to the phoretic medium container 20.

REFERENCE SIGNS LIST

10 capillary array

11 capillary

12 detecting unit

13 capillary head

14 capillary head boss

15 capillary head distal end

16 load header

17 SUS pipe

20 phoretic medium container

21 syringe

22 seal

23 rubber stopper

24 cap

26 phoretic medium

27 air

30 anode side buffer solution container

31 anode side cleaning layer

32 buffer solution layer for anode side electrophoresis

33 buffer solution layer for anode side sample introduction

35 anode side cleaning liquid

36 buffer solution for anode side electrophoresis

37 buffer solution for anode side sample introduction

40 cathode side buffer solution container

41 waste liquid layer

42 cathode side cleaning layer

43 buffer solution layer for cathode side electrophoresis

45 waste liquid received liquid

46 cathode side cleaning liquid

47 buffer solution for cathode side electrophoresis

50 sample container

51 sample

55 film

56 partition

60 liquid feeding mechanism

61 plunger

62 stepping motor

63 rotary encoder

64 origin sensor

65 ball screw

66 linear guide

67 driving pulley

68 driven pulley

69 belt

70 liquid feeding mechanism base

71 slider

72 detection plate

80 sampler base

85 Y driving member

90 Z driving member

95 X driving member

100 sample tray

101 guide

110 constant-temperature bath

115 electrode

120 constant-temperature bath door

130 irradiation detecting unit

150 auto-sampler unit

160 irradiation detecting and constant-temperature bath

200 chart of analysis workflow (each of articles of consumption set)

201 chart of analysis workflow (temperature adjusting of capillary)

202 chart of analysis workflow (cleaning of capillary)

203 chart of analysis workflow (feeding of phoretic medium)

204 chart of analysis workflow (sample introduction)

205 chart of analysis workflow (electrophoresis)

206 chart of analysis workflow (finish of analysis)

1. An electrophoresis method which is an analysis method using ananalysis device including capillaries, a capillary head that is providedat a distal end of the capillaries, a phoretic medium container that isused for electrophoresis and filled with a phoretic medium, a liquidfeeding mechanism that is provided with a plunger and feeds the phoreticmedium in the phoretic medium container to the capillaries, wherein thephoretic medium container is provided with a syringe, a seal member thatseals the phoretic medium in the syringe from below and is operatedwithin the syringe by pressing with the plunger, and a lid portion thatseals an upper part of the syringe, the method comprising: breakingthrough the lid portion with the capillary head; and pressing the lidportion using the capillary head at a time of pressing the seal memberwith the plunger.
 2. The electrophoresis method according to claim 1,further comprising: releasing a pressing force of the plunger to theseal member and pulling out the capillary head from phoretic mediumcontainer.